Registration Dossier

Environmental fate & pathways

Biodegradation in water: screening tests

Administrative data

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2007-01-11 to 2007-02-09
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report Date:
2007

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Version / remarks:
(1992)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Dr. U. NOACK-LABORATORIEN, Käthe-Paulus-Straße 1, D-31157 Sarstedt (Germany)

Test material

Reference
Name:
Unnamed
Type:
Constituent

Study design

Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Activated sludge was sampled at the municipal sewage treatment plant at Hildesheim (Germany) which treated mostly municipal sewage (industrial chemical waste: 30-33% )
- Preparation of inoculum for exposure: The activated sludge was washed twice with autoclaved tap water. After the second washing the settled sludge was filled up with mineral salts medium and was maintained in an aerobic condition by aeration for 4 h. Thereafter the sludge was homogenized with a blender. The supernatant was decanted and maintained in an aerobic condition by aeration with CO2 free air for 5 d.
- Pretreatment: no
- Initial cell/biomass concentration: 10E7-10E8 CFU/L
Duration of test (contact time):
28 d
Initial test substance concentration
Initial conc.:
20 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST CONDITIONS
- Composition of medium: according to OECD Guideline 301 B/CO2 Evolution Test
- Additional substrate: no
- Solubilising agent (type and concentration if used): not used
- Test temperature: 22 +/- 2°C
- pH: 6.61-6.76 in all test vessels on day 28
- pH adjusted: no
- Continuous darkness: no (low light conditions)

TEST SYSTEM
- Culturing apparatus: 5 L brown glass bottles
- Number of culture flasks/concentration: 2
- Method used to create aerobic conditions: The incubation vessels were aerated for 24 h with CO2 free air
- Measuring equipment: pH-Meter, Multilab 340i, WTW; Thermohygrograph, type 3.015/3 K; Flow meter, Krohne Duisburg TYP DK 800 PV
- Test performed in closed vessels: yes
- Details of trap for CO2: The outlet of the incubation vessels were connected with the adsorption vessels arranged in series. Each of the three adsorption vessels (gas wash bottles) contained 100 mL of a 0.0125 mol/L Ba(OH)2 solution. Backtitration of the residual Ba(OH)2 was carried out with 0.05 N HCl.

SAMPLING
- Sampling frequency: sampling was performed on days 1, 4, 6, 8, 11, 14, 18, 21, 25, 28, and after acidification on day 29

CONTROL AND BLANK SYSTEM
- Inoculum blank: performed
- Toxicity control: performed
- Other: functional control with the reference substance performed
Reference substance
Reference substance:
benzoic acid, sodium salt
Remarks:
(20 mg/L)

Results and discussion

% Degradation
Parameter:
% degradation (CO2 evolution)
Value:
77
Sampling time:
28 d

BOD5 / COD results

Results with reference substance:
The functional control sodium benzoate reached the pass level of 60% after 5 d and came to 100% after 14 d. The validity criterion of the guideline degradation >=60% after 14 d was fulfilled.

Any other information on results incl. tables

The biodegradation of L-tryptophan was investigated in a study conducted according to OECD Guideline 301 B. The following results were obtained for L-tryptophan, the functional control, and the toxicity control

 

Table 1: CO2 production and biodegradation in the control and test item samples

Study day

Control (mg CO2/ 3L)

Test item (20 mg/L)

Replicate 1

Replicate 2

(mg CO2/ 3L)

Degr. (%)

(mg CO2/ 3L)

Degr. (%)

Gross

Net sum

Gross

Net sum

1

4.3

3.3

0

0

3.5

0

0

4

10.7

52.9

42.2

30

52.8

42.1

30

6

10.3

46.0

77.9

55

46.9

78.7

55

8

8.5

30.0

99.4

70

28.1

98.3

69

11

10.2

20.1

109.3

77

18.2

106.3

75

14

13.7

16.2

111.8

79

13.3

106.3

75

18

14.0

14.1

111.9

79

13.0

106.3

75

21

10.3

10.5

112.1

79

8.6

106.3

75

25

12.7

12.9

112.3

79

10.8

106.3

75

28

10.3

9.8

112.3

79

9.6

106.3

75

29*

8.1

6.5

112.3

79

5.6

106.3

75

*results of the last two gas wash bottles

The test substance reached the 10% level after 2 d and the pass level of 60% after 7 d. The mean biodegradation came to 77% after 29 d. The test substance fulfills the guideline criterion of ready biodegradation and can therefore regarded as readily biodegradable.

 

Table 2: CO2 production and biodegradation in the control, functional control and toxicity control samples

Study day

Control (mg CO2/ 3L)

Functional control (20 mg/L)

Toxicity control (20 mg/L reference substance + 20 mg/L test item

(mg CO2/ 3L)

Degr. (%)

(mg CO2/ 3L)

Degr. (%)

Gross

Net sum

Gross

Net sum

1

4.3

16.3

12.0

9

3.1

0

0

4

10.7

52.4

53.7

42

53.0

42.3

16

6

10.3

46.6

90.0

70

52.8

84.8

31

8

8.5

29.8

111.3

87

52.8

129.1

48

11

10.2

21.6

122.7

96

52.8

171.7

64

14

13.7

19.7

128.7

100

45.2

203.2

75

18

14.0

22.0

136.7

100

24.5

213.7

79

21

10.3

15.1

141.5

100

20.1

223.5

83

25

12.7

17.6

146.4

100

23.7

234.5

87

28

10.3

16.5

152.6

100

17.3

241.5

89

29*

8.1

7.1

152.6

100

8.5

241.9

90

*results of the last two gas wash bottles

The functional control reached the pass level of 60% after 5 d and came to 100% after 14 d. The validity criterion degradation >=60% was fulfilled. In the toxicity control a biodegradation of 75% occurred within 14 d and came to 90% after 28 d. The biodegradation of the reference item was not inhibited by the test item. In the inoculum control a maximum of 37.7 mg CO2/L was formed after 28 d and thus fulfilling the validity criterion of the guideline (<40 mg CO2/L after 28 d). The validity criterion that the difference of extremes of replicate values of removal of the test item at the end of the test, at the plateau or at the end of the 10-d-window as appropriate must be less than 20% was also fulfilled.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
In a study conducted according to OECD Guideline 301 B, L-tryptophan proved to be readily biodegradable and fulfilling the 10-d-window criterion (77% biodegradation after 28 d).
Executive summary:

The ready biodegradation of L-tryptophan was investigated in a study conducted according to OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test) over a period of 28 days and using non-adapted activated sludge as inoculum. The biodegradation rate was determined by measurement of CO2 evolution. Inoculum blank, procedural/functional control with reference substance sodium benzoate, and toxicity control using 20 mg/L test item and 20 mg/L reference compound were performed.

 

This study is regarded as reliable without restriction and satisfies the guideline requirements for ready biodegradation.

The test item proved to be readily biodegradable and fulfilling the 10-d-window criterion (77% biodegradation after 28 d). The functional control reached the pass level > 60% after 14 d. In the toxicity control containing both test and reference item 75% biodegradation based on ThCO2 occurred within 14 d thus indicating that the test item was not inhibitory at the concentration tested.