5-nitroisophthalic acid

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

Experimental data from structurally similar read across analogue

Justification for type of information:

Experimental data from structurally similar read across analogue

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

other: refer principle below

Principles of method if other than guideline:

WoE report is prepared based on bidegradability of test chemical study: WoE-2 and WoE-3

GLP compliance:

no

Oxygen conditions:

aerobic

Inoculum or test system:

other:

Remarks:

WoE-2 Mixed inoculum and WoE-3: activated sludge, adapted

Details on inoculum:

WoE-2: Mixed Inoculum Preparation:Polyseed were used for this study. 1 polyseed capsule was added in 500 ml D.I water and then stirred for 1 hour for proper mixing and functioning of inoculum. This gave the bacterial count as 10E7 to 10E8 CFU/ml. At the regular interval microbial plating was also performed on agar to confirm the vitality and CFU count of microorganism.

WoE-3
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): sewage plant
- Laboratory culture: 1000ml volumetric cylinder
- Method of cultivation: Activated sludge taken from a sewage plant is cultivated in a 1000ml volumetric cylinder. The mixture is aerated with pressure air. Every day 200 ml of the mixture is driven off so that the sludge age is 5 days. After driving off the 200ml of the mixture aeration is interrupted, and after sedimentation ca. 600ml of the liquid phase is driven off. The residue (200 ml of the thickened activated sludge) is diluted with tap water to the volume of ca. 800 ml and 600 mg/l of starch or glucose, 600 mg/l of peptone, 25 ml of a phosphate buffer pH 7.2, and the solution of the tested compound are added. Then the mixture in the cylinder is made up to 1000ml with tap water and aerated for 23 h (the recirculation ratio is 0-25). After this period the procedure is repeated.
- Initial cell/biomass concentration: 100 mg/l

Duration of test (contact time):

28 d

Initial conc.:

4 mg/L

Based on:

test mat.

Initial conc.:

200 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Parameter followed for biodegradation estimation:

other: COD removal

Details on study design:

WoE-2:

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 BOD bottles; with glass stoppers (125 ml), BOD incubator & oxygen electrode and meter.

CONTROL AND BLANK SYSTEM
- Inoculum blank: it contains only test inoculum
- Procedure control: contains reference compound and inoculum


WoE-3
TEST CONDITIONS
- Composition of medium: In ca. 800 ml of distilled water solutions of calcium chloride (27.5 g CaCl2 in 1l Distilled water) magnesium sulphate (22.5 g MgSO4.7H20 in I 1. distilled water), and ferric chloride (0.25 g FeCI3.6 H20 in 11. distilled water) are added in 1 ml portions each. Then 5 ml ammonium sulphate solution [10g (NH4)2SO4 in 11. distilled water], 20ml of a phosphate buffer of pH 7.2 (8.5 g KH2PO4, 21.8g K2HPO4, and 44.7g Na2HPO4. 12 H2O in 1l distilled water), and 100ml of tap water for securing the content of trace elements, are added. The solution thus prepared is made up to 1000ml with distilled water.
- Test temperature: 20±3°C
- pH: 7.2
- pH adjusted: No
- Aeration of dilution water: The mixture is aerated with pressure air
- Continuous darkness: Yes , dark room
- Other: Beaker containing inoculums is placed on electromagnetic stirrer.

CONTROL AND BLANK SYSTEM
Inoculum blank: Yes

Reference substance:

benzoic acid, sodium salt

Key result

Parameter:

% degradation (O2 consumption)

Value:

73.33

Sampling time:

28 d

Remarks on result:

other: Other details not known

Parameter:

other: COD removal

Value:

96.8

Sampling time:

20 d

Remarks on result:

other: Sampling time - < 20 days

Details on results:

WoE-2:
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 0.77 mgO2/mg. ThOD was determined by calculation as 1.05 mgO2/mg. % degradation was calculated using the values of BOD and ThOD for test item and was determined to be 73.33% at 20 ± 1°C. The % degradation of procedure control (reference item) was also calculated using BOD & ThOD and was determined to be 75.3%. Degradation of Sodium Benzoate exceeds 46.38% on 7 days & 61.44% on 14th day. The activity of the inoculum was thus verified and the test can be considered as valid.

WoE-3
The percentage degradation of test chemical was determined to be 94.6% in < 20 days by using COD removal parameter.

Results with reference substance:

WoE-2:
The % degradation of procedure control (reference item) was also calculated using BOD & ThOD and was determined to be 75.3%. Degradation of Sodium Benzoate exceeds 46.38% on 7 days & 61.44% on 14th day. The activity of the inoculum was thus verified and the test can be considered as valid.

WoE-2:

TABLE 2: D.O Values (mg/L)

No. of Days	Inoculum Blank (Control)	Test Suspension	Procedure Control (Reference Item)
0	7.4	7.3	7.2
7	6.9	5.4	3.6
14	6.6	4	2.3
21	6.2	3.4	1.5
28	5.9	2.7	0.7

TABLE 2: BOD Values (mg O2/mg)

 

No. of Days	Test Suspension	Procedure Control (Reference Item)
0	0	0
7	0.35	0.77
14	0.62	1.02
21	0.67	1.12
28	0.77	1.25

 

TABLE 3: PERCENT BIODEGRDATION RESULTS

No. of Days	Test Suspension	Procedure Control (Reference Item)
0	0%	0%
7	33.33%	46.38%
14	59.04%	61.44%
21	65.8%	67.46%
28	73.33%	75.3%

 

Table 4: BOD28, THOD AND % BIODEGRADATION VALUES

 

Method details	BOD28 (mgO2/mg)	ThOD (mgO2/mg)	%Biodegradation
Test Item	0.77	1.05	73.33
Reference Item	1.25	1.66	75.3

WoE-3

Table: Biodegradability of the test compounds

 

 

Compound	Percent removed(based upon COD)	Rate of biodegradation (mg COD g-1h-1)
Test chemical	96.8	78.4

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable

Conclusions:

By applying weight of evidence approach the percent biodegradation of test chemical is expected to be in range from 73.33% to 96.8% in aerobic condition and in 28 days. On the basis of this percent biodegradability test chemical is considered to be readily biodegradable.

Executive summary:

Biodegradability of test chemical was determined by using weight of evidence approach by using different experimental data from its structurally similar read across analogues and their results are summarized below

 

In first study the Closed Bottle test following the OECD guideline 301 D was conducted to determine the ready biodegradability of the test chemical. The study was performed at a temperature of 20°C. The test system included control, test chemical and reference chemical. Polyseed were used as inoculum. The concentration of test and reference chemical (Sodium Benzoate) chosen for both the study was 4 mg/L, while that of inoculum was 32 ml/l. OECD mineral medium was used for the study. ThOD (Theoretical oxygen demand) of test and reference chemical was determined by calculation. % degradation was calculated using the values of BOD and ThOD for test chemical and reference chemical. The % degradation of procedure control (reference chemical) was also calculated using BOD & ThOD and was determined to be 75.3%. Degradation of Sodium Benzoate exceeds 46.38% on 7 days & 61.44% on 14th day. The activity of the inoculum was thus verified and the test can be considered as valid.The BOD28 value of test chemical was observed to be 0.77 mgO2/mg. ThOD was calculated as 1.05 mgO2/mg. Accordingly, the % degradation of the test chemical after 28 days of incubation at 20 ± 1°C according to Closed Bottle test was determined to be 73.33%. Based on the results, the test chemical, under the test conditions, was considered to be readily biodegradable in nature.

 

Another study was reviewed from journal Water Research, 1976 in this a batch test in an open system was conducted for 20 days for evaluating the biodegradability of test chemical. Adapted activated sludge was used as a test inoculum obtained from a sewage plant is cultivated in a1000ml volumetric cylinder. The mixture is aerated with pressure air. Every day 200 ml of the mixture is driven off so that the sludge age is 5 days. After driving off the 200ml of the mixture aeration is interrupted, and after sedimentation ca.600ml of the liquid phase is driven off. The residue (200 ml of the thickened activated sludge) is diluted with tap water to the volume ofca.800 ml and 600 mg/l of starch or glucose, 600 mg/l of peptone, 25 ml of a phosphate buffer pH 7.2, and the solution of the tested compound are added. Then the mixture in the cylinder is made up to 1000ml with tap water and aerated for 23 h (the recirculation ratio is 0-25). After this period the procedure is repeated. Test chemical conc. used for the study was 200 mg/l based on COD. To 1000-1500ml of the biological medium such amount of the solution of the substance tested is added that the initial COD is 200 mg/l. Then such an amount of the adapted activated sludge, washed and thickened by sedimentation, is dosed to the medium that the concentration of the dry matter is 100 mg/l. simultaneously, a blank test is prepared. The beaker is placed in a dark room with a roughly 3 constant temperature of 20±3°C on an electromagnetic stirrer and a pH of 7.2 for 120 hrs. The initial value of COD or organic carbon of the liquid phase are determined. Samples filtered or centrifuged before analysis, are taken at suitable intervals. The decrease of the tested substance in the liquid phase is evaluated by determining COD or organic carbon. The results are compared with those of a blank test and standard compound decomposition. With the degree of degradation also the average specific rate of degradation is determined, expressed in terms of mg COD (or organic carbon) removed by a gram of dry matter of the activated sludge per hour. The percentage degradation of test chemical was determined to be 96.8% by using COD removal parameter in < 20 days. Thus, based on percentage degradation, test chemical was considered to be readily biodegradable in nature.

 

By considering results of both the studies mentioned above and applying weight of evidence approach the percent biodegradation of test chemical is expected to be in range from 73.33% to 96.8% in aerobic condition and in 28 days. On the basis of this percent biodegradability test chemical is considered to be readily biodegradable.

Description of key information

By applying weight of evidence approach the percent biodegradation of test chemical is expected to be in range from 73.33% to 96.8% in aerobic condition and in 28 days. On the basis of this percent biodegradability test chemical is considered to be readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Type of water:

freshwater

Additional information

Biodegradability of test chemical was determined by using weight of evidence approach by using different experimental data from its structurally similar read across analogues and their results are summarized below

 

In first study the Closed Bottle test following the OECD guideline 301 D was conducted to determine the ready biodegradability of the test chemical. The study was performed at a temperature of 20°C. The test system included control, test chemical and reference chemical. Polyseed were used as inoculum. The concentration of test and reference chemical (Sodium Benzoate) chosen for both the study was 4 mg/L, while that of inoculum was 32 ml/l. OECD mineral medium was used for the study. ThOD (Theoretical oxygen demand) of test and reference chemical was determined by calculation. % degradation was calculated using the values of BOD and ThOD for test chemical and reference chemical. The % degradation of procedure control (reference chemical) was also calculated using BOD & ThOD and was determined to be 75.3%. Degradation of Sodium Benzoate exceeds 46.38% on 7 days & 61.44% on 14th day. The activity of the inoculum was thus verified and the test can be considered as valid.The BOD28 value of test chemical was observed to be 0.77 mgO2/mg. ThOD was calculated as 1.05 mgO2/mg. Accordingly, the % degradation of the test chemical after 28 days of incubation at 20 ± 1°C according to Closed Bottle test was determined to be 73.33%. Based on the results, the test chemical, under the test conditions, was considered to be readily biodegradable in nature.

 

Another study was reviewed from journal Water Research, 1976 in this a batch test in an open system was conducted for 20 days for evaluating the biodegradability of test chemical. Adapted activated sludge was used as a test inoculum obtained from a sewage plant is cultivated in a1000ml volumetric cylinder. The mixture is aerated with pressure air. Every day 200 ml of the mixture is driven off so that the sludge age is 5 days. After driving off the 200ml of the mixture aeration is interrupted, and after sedimentation ca.600ml of the liquid phase is driven off. The residue (200 ml of the thickened activated sludge) is diluted with tap water to the volume ofca.800 ml and 600 mg/l of starch or glucose, 600 mg/l of peptone, 25 ml of a phosphate buffer pH 7.2, and the solution of the tested compound are added. Then the mixture in the cylinder is made up to 1000ml with tap water and aerated for 23 h (the recirculation ratio is 0-25). After this period the procedure is repeated. Test chemical conc. used for the study was 200 mg/l based on COD. To 1000-1500ml of the biological medium such amount of the solution of the substance tested is added that the initial COD is 200 mg/l. Then such an amount of the adapted activated sludge, washed and thickened by sedimentation, is dosed to the medium that the concentration of the dry matter is 100 mg/l. simultaneously, a blank test is prepared. The beaker is placed in a dark room with a roughly 3 constant temperature of 20±3°C on an electromagnetic stirrer and a pH of 7.2 for 120 hrs. The initial value of COD or organic carbon of the liquid phase are determined. Samples filtered or centrifuged before analysis, are taken at suitable intervals. The decrease of the tested substance in the liquid phase is evaluated by determining COD or organic carbon. The results are compared with those of a blank test and standard compound decomposition. With the degree of degradation also the average specific rate of degradation is determined, expressed in terms of mg COD (or organic carbon) removed by a gram of dry matter of the activated sludge per hour. The percentage degradation of test chemical was determined to be 96.8% by using COD removal parameter in < 20 days. Thus, based on percentage degradation, test chemical was considered to be readily biodegradable in nature.

 

By considering results of both the studies mentioned above and applying weight of evidence approach the percent biodegradation of test chemical is expected to be in range from 73.33% to 96.8% in aerobic condition and in 28 days. On the basis of this percent biodegradability test chemical is considered to be readily biodegradable.