Ethyl diphenylcarbamate

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

November 8, 2016 - December 6, 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: Aeration tank of Sewage Treatment Plant "Czajka", Warsaw, receiving predominantly domestic sludge.
- Preparation of inoculum for exposure: The coarse particles were removed by settling and the supernatant was discarded. The sludge was washed in the mineral medium. The concentrated sludge was suspended in mineral medium to yield a concentration of 3-5 g suspended solids/l and it was aerated for 5 days, at the test temperature of 22ºC, until application. A sample was withdrawn just before use for determination of the dry weight of the suspended solids.
- Pretreatment: Inocula was pre-conditioned to the experimental conditions. Pre-conditioning consisted of aerating activated sludge in mineral medium at the test temperature of 22 ºC.
- Concentration of sludge: 30 mg/L

Duration of test (contact time):

ca. 28 d

Initial conc.:

ca. 100 mg/L

Based on:

act. ingr.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium: (a) Monopotassium dihydrogen orthophosphate, KH2PO4 8.50 g / Dipotassium monohydrogen orthophosphate, K2HPO4 21.75 g / Disodium monohydrogen orthophosphate dihydrate Na2HPO4 · 2 H2O 33.40 g / Ammonium chloride, NH4Cl 0.50 g. Dissolve in water and make up to 1 litre The pH of the solution should be 7.4.
(b) Calcium chloride, anhydrous, CaCl2 27.50 g. Dissolve in water and make up to 1 litre
(c) Magnesium sulphate heptahydrate, MgSO4 · 7 H2O 22.50 g. Dissolve in water and make up to 1 litre. (d) Iron(III) chloride hexahydrate, FeC13 · 6H2O 0.25 g; Dissolve in water and make up to 1 litre. Mix 10 ml of solution (a) with 800 ml dilution water, add 1 ml of solutions (b) to (d) and make up to 11 with dilution water.
- Solubilising agent: Double-distilled water containing 4 mg/L of organic carbon (<10% of the organic carbon content introduced by the test material).
- Test temperature: 22 ºC ± 2°C
- pH: 7.4 ± 0.2
- pH adjusted: no
- Suspended solids concentration: 30 mg/l
- Continuous darkness: yes
- Flasks volume: 0.164 L

TEST SYSTEM
- Culturing apparatus: Thermo-cabinet WTW TS 606 CZ-G/3-VAR. The temperature in thermo-cabinet was recorded with temperature recorder EBI 310 –T from
Ebro.
- Number of culture flasks/concentration: Duplicates, flasks#17 and 18 containing test suspension containing test item (100 mg/L) and inoculum 30 mg/L SS.
- Measuring equipment: Closed WTW OxiTop OC 110 respirometer for BOD determination

SAMPLING
- Sampling frequency: The data were read out every 112 min during the 28 day test (40320 min that is 360 readings) and were recoded and stored in the measuring heads of the sample bottles.

CONTROL AND BLANK SYSTEM
- Inoculum blank: Triplicates, flasks#1, 2 and 3 containing only inoculum 30 mg/L SS.
- Procedure control: Triplicates, flasks#4, 5 and 6 containing reference item (sodium acetate 100 mg/L) and inoculum 30 mg/SS
- Toxicity control: Duplicates, flasks#19 and 20 containing test item, reference item at the same concentrations as in the individual solutions and inoculum 30 mg/L SS.

STATISTICAL METHODS:
The calculations and the graphs were performed using SigmaPlot 9.0 software of SYSTAT Software, Inc., USA purchased from GAMBIT CoiS Ltd, Poland.

Reference substance:

acetic acid, sodium salt

Remarks:

CAS:127-09-3, pure p.a min 98.5, Eurochem

Test performance:

In this test item as a nitrogen-containing substance may affect the oxygen uptake because of nitrification reaction. The observed oxygen uptake by the reaction mixture may be corrected for the amount of oxygen used in oxidizing ammonium to nitrite and nitrate.

Key result

Parameter:

% degradation (O2 consumption)

Value:

ca. 2.3

Sampling time:

28 d

Details on results:

At the 28th day of the test the aerobic biodegradation of the testing, the test item attained 2.3% of biodegradation. The pH values of all flasks were inside the range 7.19-8.61.
In the toxicity test the biodegradation after 28 days was 18.9%, which is less than 25% (based on ThOD) occurred in 14 days. Therefore, the test item is inhibitory.
The oxigen uptake of the inoculum blank was equal to 36.9 mg/L in 28 days.

Results with reference substance:

The reference item reached 96.1% of biodegradation at the 28th day of the test.

The mineral medium solution pH was equal to 7.06.

The preconditioned suspension of inoculum pH was equal to 7.27.

The initial concentration of test item in test flasks was equal to 100 mg/L.

 

Table 1. Correction for oxygen uptake for interference by nitrification

days	0		28		difference
1) Concentration of nitrate (mg N-NO3/l)	#19	#20	#19	#20	#19	#20
	0.306	0.316	0.320	0.328	0.014	0.012
					0.013±0.001
2) Oxigen equivalent (4.57 x N-NO3) (mg/L)					0.059
3) Concentration of nitrite (mg N-NO2/l)	#19	#20	#19	#20	#19	#20
	0.016	0.017	0.015	0.016	-0.001	-0.001
					-0.001±0.0000
4) Oxigen equivalent (3.43 x N-NO2) (mg/L)					-0.003
5) total oxigen equivalent 2) + 4)					0.056

 

Toxicity test

days	0		28		difference
1) Concentration of nitrate (mg N-NO3/l)	#19	#20	#19	#20	#19	#20
	0.278	0.272	0.281	0.279	0.003	0.007
					0.005±0.002
2) Oxigen equivalent (4.57 x N-NO3) (mg/L)					0.023
3) Concentration of nitrite (mg N-NO2/l)	#19	#20	#19	#20	#19	#20
	0.017	0.016	0.015	0.015	-0.002	-0.001
					0.0015±0.0005
4) Oxigen equivalent (3.43 x N-NO2) (mg/L)					-0.005
5) total oxigen equivalent 2) + 4)					0.018

 

Table 2. The pH values of test flasks

Flask #	17	18	1	2	3	4	5	6	19	20
	Test item		Control			Reference item			Toxicity test
Initial	7.59	7.60	7.44	7.44	7.45	7.43	7.45	7.44	7.58	7.56
final	7.52	7.67	7.21	7.19	7.25	8.22	8.11	8.33	8.61	8.47

No adjustment of pH was conducted

Table 3. Sample oxygen uptake: biodegradability

		Time, days
		1	3	5	7	9	12	14	16	18	21	23	25	28
Test item O2uptake, mg/L	a1	0.0	0.0	3.1	4.6	6.2	8.4	12.1	17.7	22.5	28.3	31.2	34.2	36.6
	a2	0.0	3.1	7.8	14.7	21.0	29.4	32.4	36.4	36.6	40.5	42.2	45.0	47.8
	amavg	0.0	1.5	5.4	9.7	13.6	18.9	22.2	27.1	29.5	34.4	36.7	39.6	42.2
Blank test O2uptake, mg/L	b1	4.8	14.4	17.4	21.0	22.3	27.3	29.3	30.1	34.3	37.0	38.2	39.8	44.1
	b2	3.6	8.7	14.7	16.4	22.2	22.9	24.7	26.2	25.9	25.3	29.2	29.1	31.1
	b3	3.6	14.0	18.0	21.7	25.0	27.2	28.3	28.0	33.2	32.5	32.4	34.1	35.5
	bmavg	4.0	12.4	16.7	19.7	23.1	25.8	27.5	28.1	31.2	31.6	33.3	34.4	36.9
Reference item O2uptake, mg/L	w1	19.1	53.8	67.1	76.5	84.5	92.3	95.7	98.6	101.2	106.2	108.0	110.8	116.2
	w2	21.7	52.2	62.4	73.1	79.9	87.3	92.6	96.5	98.9	104.5	106.2	109.7	113.6
	w3	20.9	50.7	63.1	70.5	78.4	85.2	89.3	92.8	94.4	99.2	103.3	105.8	110.1
	wmavg	20.5	52.2	64.2	73.3	80.9	88.3	92.5	95.9	98.2	103.3	105.8	108.7	113.3
Toxicity control O2uptake, mg/L	a4tox1	0.0	52.5	67.8	74.0	79.5	92.9	102.3	115.1	124.0	135.4	141.8	146.2	155.4
	a5tox2	1.7	53.0	62.5	70.0	73.2	78.2	81.6	83.5	87.9	90.0	90.3	93.1	94.3
	toxmavg	1.7	53.0	62.5	70.0	73.2	78.2	81.6	83.5	87.9	90.0	90.3	93.1	94.3
Corrected test item O2uptake, mg/L	a1- bm	-4.5	-12.4	-13.6	-15.2	-16.9	-17.4	-15.4	-10.4	-8.7	-3.3	-2.1	-0.2	-0.3
	a2- bm	-4.0	-9.3	-8.9	-5.0	-2.2	3.5	4.9	8.3	5.4	8.9	8.9	10.6	10.9
Reference item % biodegradation (BOD/ThODxC)x100   ThOD=0.78 mgO2/mg C=100 mg/L	R1(w1)	19.4	53.1	64.6	72.8	78.7	85.3	87.5	90.3	89.9	95.6	95.8	98.0	101.6
	R2(w2)	22.6	51.1	58.7	68.4	72.8	78.9	83.5	87.6	86.8	93.4	93.5	96.6	98.4
	R3(w3)	21.6	49.2	59.5	65.1	70.8	76.1	79.3	82.9	81.1	86.7	89.7	91.6	93.8
	Rtoxavg	22.1	50.1	59.1	66.7	71.8	77.5	81.4	85.2	84.0	90.0	91.6	94.1	96.1

The reference item biodegradability result R₁(w1) is not taken into average value calculations, as it is greater than 100% on 28^thday. The toxicity control result a_4tox1 was rejected as erroneously high.

Validity criteria fulfilled:

yes

Remarks:

(difference of replicates <20%,reference item reached pass level (60%) on day 5, biodegradation of toxicity test on day 28 was 95.3%,oxigen uptake of blank was 36.9 mg O2/l in 28 days,pH values were inside 6-8.5 range,oxigen consumptiom by test item <60%)

Interpretation of results:

not readily biodegradable

Conclusions:

At the 28th day, the test item attained 2.3% of biodegradation, and it can be assumed to be inhibitory. Therefore, the test item cannot be deemed to be readily biodegradable.

Executive summary:

The study of ready biodegradability of test substance in an aerobic aqueous medium with manometric respirometry method was performed according to OECD 301F and EU C.4 -D method, following GLP. 100 mg/l of test item was inoculated with activated sludge (30 mg/L SS) and incubated under aerobic conditions in a closed respirometer flask at constant temperature (22 ± 2ºC) for 28 days. The observed oxygen uptake by the reaction mixture was corrected for the amount of oxygen used in oxidising ammonium to nitrite and nitrate. All the acceptability criteria were met and the study was considered valid. A blank test, a procedure test with reference substance (sodium acetate) and a toxicity test were run in parallel. At the 28 th day, the reference item reached 96.1% of biodegradation. In a toxicity test, containing both the test item and a reference item, on the 28th day the biodegradation was 18.9%. Thus, the test item can be assumed to be inhibitory. At the 28th day of the test the measured aerobic biodegradation of the test item equals 2.3%. Therefore, the test item cannot be deemed to be readily biodegradable.

Description of key information

Key study: OECD 301F and EU C.4 - D. GLP study. At the 28th day, the test item attained 2.3% of biodegradation, and it can be assumed to be inhibitory. Therefore, the test item cannot be deemed to be readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Additional information

Key study: The study of ready biodegradability of test substance in an aerobic aqueous medium with manometric respirometry method was performed according to OECD 301F and EU C.4 -D method, following GLP. 100 mg/l of test item was inoculated with activated sludge (30 mg/L SS) and incubated under aerobic conditions in a closed respirometer flask at constant temperature (22 ± 2ºC) for 28 days. The observed oxygen uptake by the reaction mixture was corrected for the amount of oxygen used in oxidising ammonium to nitrite and nitrate. All the acceptability criteria were met and the study was considered valid. A blank test, a procedure test with reference substance (sodium acetate) and a toxicity test were run in parallel. At the 28 th day, the reference item reached 96.1% of biodegradation. In a toxicity test, containing both the test item and a reference item, on the 28th day the biodegradation was 18.9%. Thus, the test item can be assumed to be inhibitory. At the 28th day of the test the measured aerobic biodegradation of the test item equals 2.3%. Therefore, the test item cannot be deemed to be readily biodegradable.