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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Experimental result using OECD guidelines.
Qualifier:
according to guideline
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Principles of method if other than guideline:
Biodegradation study was conducted according to OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test) for evaluating the percentage biodegradability of test chemical 2,4,6-trichloroaniline
GLP compliance:
no
Specific details on test material used for the study:
-Name of the test material: 2,4,6-trichloroaniline
- Molecular formula: C6H4Cl3N
- Molecular weight: 196.464 g/mol
- Smiles notation: Nc1c(Cl)cc(Cl)cc1Cl
- InChl : 1/C6H4Cl3N/c7-3-1-4(8)6(10)5(9)2-3/h1-2H,10H2
-Substance type: Organic
-Physical state: solid
Oxygen conditions:
aerobic
Inoculum or test system:
other: mixture of domestic waste water, surface soil and soil samples
Details on inoculum:
Mixed Inoculum Preparation (In-House): Domestic waste water, surface soil and soil samples from polluted sites receiving predominantly domestic waste was collected and were mixed to get diluted suspension. The inoculum was kept aerobic until being used for experiment by supplying organic and inorganic sources required by micro flora to sustain at controlled laboratory conditions. This gave the bacterial count as 107to 108 CFU/ml. At the regular interval microbial plating was also performed on agar to confirm the vitality and CFU count of microorganism.
Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
other: BOD and ThOD
Details on study design:
TEST CONDITIONS
- Composition of medium: OECD mineral medium was used for the study
- Test temperature: 20°C
- Continuous darkness: Yes
- Other: The water used in this study is deionized water.

TEST SYSTEM
- Culturing apparatus: The apparatus used in this study is Respirometer BOD Sensor System (Code: F102B0133; non-automatic pressure measurement sensor system) and Respirometric incubator (FOC 120i)) by VELP Scientifica.

CONTROL AND BLANK SYSTEM
- Inoculum blank: it contains only test inoculum
- Procedure control: contains reference compound and inoculum
Reference substance:
other: Sodium Benzoate
Parameter:
other: BOD and ThOD
Value:
2.57
Sampling time:
28 d
Remarks on result:
other: Other details not known
Details on results:
The oxygen consumed by the test systems was corrected for oxygen consumption occurring in the blank test systems. The BOD Values (mgO2/mg) and percent biodegradation results for each test system are reported in Tables 2 and 3, respectively. The BOD28 value of test chemical was observed to be 0.023 mgO2/mg. ThoD was determined by calculation as 0.896 mgO2/mg. % Degradation was calculated using the values of BOD and ThOD for test item and was dertermined to be 2.57 %. The % degradation of procedure control (Sodium Benzoate) was also calculated using BOD & ThOD and was found to be 74.074%. The mean BOD values (mg O2/l) in control on 28th day was 9 mg O2/l, thereby fulfilling the control validity criteria (i.e., the oxygen consumption of the inoculum blank is normally 20-30 mg O2/l and should not be greater than 60 mg/l in 28 days). Degradation of Sodium Benzoate exceeds 50.050 %after 7 days and 68.068 % after 14 days. The activity of the inoculums is thus verified and the test can be considered as valid.
Results with reference substance:
The % degradation of procedure control (Sodium Benzoate) was also calculated using BOD & ThOD and was found to be 75.20%. The mean BOD values (mg O2/l) in control on 28th day was 1.25 mg O2/l, thereby fulfilling the control validity criteria (i.e., The oxygen consumption of the inoculum blank is normally 20-30 mg O2/l and should not be greater than 60 mg/l in 28 days). Degradation of Sodium Benzoate exceeds 65.65 %after 7 days and 69.79 % after 14 days. The activity of the inoculums is thus verified and the test can be considered as valid.

PERCENT BIODEGRADATION RESULTS

No. of Days

Test Suspension

Procedure Control (Reference Item)

1

-2.46

15.68

2

-1.23

38.02

3

-1.23

52.73

4

-0.56

58.74

5

0.11

61.14

6

1.23

65.05

7

0.67

65.65

8

0.67

66.25

9

0.67

67.75

10

0.67

68.35

11

0.11

68.35

12

0.67

69.85

13

1.34

70.15

14

0.67

69.79

15

1.23

70.69

16

1.34

71.95

17

1.34

72.55

18

1.34

71.89

19

1.34

71.89

20

1.90

72.19

21

1.90

73.39

22

2.57

73.09

23

1.90

73.69

24

2.57

74.29

25

1.90

74.29

26

2.57

74.29

27

2.57

74.29

28

2.57

75.20

BOD28, ThOD AND % BIODEGRADATION VALUES

Method details

BOD28(mgO2/mg)

ThOD (mgO2/mg)

% Biodegradation

Test Item

0.023

0.896

2.57

Reference Item

1.25

1.665

75.20

Validity criteria fulfilled:
yes
Interpretation of results:
not readily biodegradable
Conclusions:
The test chemical 2,4,6-trichloroaniline undergoes 2.57 % biodegradation after 28 days in the test condition. Thus, the test chemical can be considered as not readily biodegradable.
Executive summary:

28-days Manometric respirometry test following the OECD guideline 301F to determine the ready biodegradability of the test chemical 2,4,6-trichloroaniline . The study was performed at a temperature of 20± 1°C. Mixture of domestic waste water, surface soil and soil samples was used as a test inoculum for the study. This inoculum was collected and were mixed to get diluted suspension. The inoculum was kept aerobic until being used for experiment by supplying organic and inorganic sources required by micro flora to sustain at controlled laboratory conditions.  This gave the bacterial count as 107 to 108 CFU/ml. At the regular interval microbial plating was also performed on agar to confirm the vitality and CFU count of microorganism. OECD mineral medium was used for the study. The test system included control, test item and reference item. The concentration of test and reference item ( Sodium Benzoate) chosen for both the study was 100 mg/L, while that of inoculum was 10 ml/L . ThOD (Theoretical oxygen demand) of test and reference item was determined by calculation. % Degradation was calculated using the values of BOD and ThOD for test item and reference item. The % degradation of procedure control (Sodium Benzoate) was also calculated using BOD & ThOD and was determined to be 75.20%. The mean BOD values (mg O2/l) in control on 28th day was 15.8mg O2/l, thereby fulfilling the control validity criteria (i.e., The oxygen consumption of the inoculum blank is normally 20-30 mg O2/l and should not be greater than 60 mg/l in 28 days). Degradation of Sodium Benzoate exceeds 65.65 %after 7 days and 69.79 % after 14 days. The activity of the inoculums is thus verified and the test can be considered as valid. The BOD28 value of test chemical   2,4,6-trichloroaniline was observed to be 0.023 mgO2sup>/mg. ThOD was calculated as 0.896 mgO2/mg. Accordingly, the % degradation of the test item after 28 days of incubation at 20 ± 1°C according to manometric respirometry test was determined to be 2.57 %.Based on the results, the test item, under the test conditions, was considered to be not readily biodegradable at 20 ± 1°C over a period of 28 days.

Description of key information

The test chemical 2,4,6-trichloroaniline undergoes 2.57 % biodegradation after 28 days in the test condition. Thus, the test chemical can be considered as not readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:
under test conditions no biodegradation observed

Additional information

Various experimental studies were reviewed from experimental report and different journals to determine biodegradability of test chemical 2,4,6-trichloroaniline in water and their results are summarized below.

 

The first experimental study was reviewed from experimental report in this study the 28-days Manometric respirometry test following the OECD guideline 301F was performed to determine the ready biodegradability of the test chemical 2,4,6-trichloroaniline . The study was performed at a temperature of 20± 1°C. Mixture of domestic waste water, surface soil and soil samples was used as a test inoculum for the study. This inoculum was collected and was mixed to get diluted suspension. The inoculum was kept aerobic until being used for experiment by supplying organic and inorganic sources required by micro flora to sustain at controlled laboratory conditions.  This gave the bacterial count 107 to 108 CFU/ml. At the regular interval microbial plating was also performed on agar to confirm the vitality and CFU count of microorganism. OECD mineral medium was used for the study. The test system included control, test item and reference item. The concentration of test and reference item (Sodium Benzoate) chosen for both the study was 100 mg/L, while that of inoculum was 10 ml/L . ThOD (Theoretical oxygen demand) of test and reference item was determined by calculation. % Degradation was calculated using the values of BOD and ThOD for test item and reference item. The % degradation of procedure control (Sodium Benzoate) was also calculated using BOD & ThOD and was determined to be 75.20%. The mean BOD value (mg O2/l) in control on 28th day was 15.8mg O2/l, thereby fulfilling the control validity criteria (i.e., the oxygen consumption of the inoculum blank is normally 20-30 mg O2/l and should not be greater than 60 mg/l in 28 days). Degradation of Sodium Benzoate exceeds 65.65 %after 7 days and 69.79 % after 14 days. The activity of the inoculums is thus verified and the test can be considered as valid. The BOD28 value of test chemical 2,4,6-trichloroanilinewas observed to be 0.023 mgO2sup>/mg. ThOD was calculated as 0.896 mgO2/mg. Accordingly, the % degradation of the test item after 28 days of incubation at 20 ± 1°C according to manometric respirometry test was determined to be 2.57 %.Based on the results, the test item, under the test conditions, was considered to be not readily biodegradable at 20 ± 1°C over a period of 28 days.

 

Next study was reviewed from Journal of Environmental Science and Health (1984) in this study Microbial degradation of test chemical 2,4,6-trichloroaniline (CAS no. 634-93-5) was conducted with water samples from which sediments were removed by settling and filtration and it is obtained from canal creek. Canal Creek is a small stream at the Aberdeen Proving Ground, MD, the sediments of which were believed to be contaminated with2,4,6-trichloroaniline.Cotton-stoppered 500 mL Erlenmeyer flasks containing 200 mL of sample water were buffered (0.2% potassium phosphate, 0.01% ammonium sulfate, pH 7.0) and incubated with 3 mg/L2,4,6-trichloroanilinein a final volume of 250 mL. Incubation was carried out in the dark for 6 weeks at 25°C with constant agitation. Aliquots of samples were removed throughout the incubation period for chemical analysis as described below or for bacterial enumeration by triplicate plating on Standard Methods Agar (SMA). Controls consisted of identical flasks containing water samples which had been autoclaved for 15minutes prior to the addition of the t e s t chemical and buffer. Parameter used for biodegradability was test material analysis by GC. More than 65 % percent degradation obtained in both control and test flask till 30 days and test chemical was 100% (beyond detection limit) in 35 days. Since degradation obtained in both control and test flask it cannot be clearly interpreted that test chemical is biodegradable or not but after 30 days test chemical is degraded beyond detection limit. On the basis of percent degradability in control as well as in test flask it can be interpreted that test chemical is not readily biodegradable.

 

Another experimental study was reviewed from authoritative database (J check, 2018) in this study the Biodegradation experiment was carried out of test chemical 2,4,6-trichloroaniline (CAS no. 634-93-5) by taking activated sludge as inoculums at 30 mg/L concentration for 28 days. Biodegradation was analyzed by using two parameters namely BOD and HPLC. The initial concentration of test chemical was 100 mg/L. After 28 days of incubation percent biodegradation of test chemical 2,4,6-trichloroaniline was observed to be 0.0 % by BOD parameter 3.0 % by GC parameter. Therefore it is concluded that test chemical 2,4,6-trichloroaniline is not readily biodegradable.

 

Last experimental study was reviewed from chemosphere journal (1985) in this study the percent biodegradation of test chemical 2,4,6-trichloroaniline (CAS no. 634-93-5) was determined by using activated sludge as inoculums. The activated sludge was obtained from municipal sewage treatment plant and it is adapted to a synthetic nutrient medium for easy handling and standardization, CO2 evolution was used as parameter. The percent biodegradation of test chemical 2,4,6-trichloroaniline was determined to be 0.4 % .On the basis of percent degradation value it can be concluded that test chemical is not readily biodegradable in nature.

 

 By considering results of all the studies mentioned above it can be concluded that test chemical 2,4,6-trichloroaniline (CAS no. 634-93-5) is not readily biodegradable in water.